The present invention is directed to polyvalent immunizing preparations composed of nontoxic serotype-specific polysaccharide preparations. More particularly, the invention described herein relates to the treatment of purified capsular polysaccharides (CPS) derived from Klebsiella species bacilli, hereinafter referred to as Klebsiella, with a deacylating agent such as sodium hydroxide (NaOH) to yield a nonpyrogenic, immunogenic CPS preparation. The present invention also relates to the use of immunogenic Klebsiella species CPS to provide protection against Klebsiella infections. A vaccine formulated from such antigens can be utilized to actively immunize individuals at risk to Klebsiella infections.
The frequency of infections due to gram-negative aerobic bacilli, such as Klebsiella, has increased dramatically over the preceding three decades, concommitant with the widespread use of broad spectrum antibiotics. Of these infections, those classified as pneumonia or bacteremia carry the highest attendant fatality ratios which average roughly 25% and are in the range of 10% to 50%. As described in McGowan, J. E., Barnes, M. W., Finland, M., "Bacteremia at Boston City Hospital: occurrence and mortality during 12 selected years (1935-1972), with special reference to hospital acquired cases," J. Infect. Dis. 132: 316-335, 1975; Bryan C. S., Reynolds, K. L., Brenner, E. R., "Analysis of 1,186 episodes of gram-negative bacteremia in non-university hospitals: the effects of antimicrobial therapy," Rev. Infect. Dis. 5: 629-638 1983; Graybill, J. R., Marshall, L. W., Charache, P., Wallace, C. K., Melvin, V. B., "Nosocomial pneumonia. A continuing major problem," Am. Rev. Resp. Dis. 108: 1130-1140, 1973; and Cross, A., Allen, J. R., Burke, J., Ducel, G., Harris, A., John, J., Johnson, D., Lew, M., MacMillan, B., Meers, P., Skalova, R., Wenzel, R., Tenney, J., "Nosocomial Infections due to Pseudomonas aeruginosa: review of recent trends," Rev. Infect. Dis. 5 (Suppl.): S837-S845, 1983, Klebsiella is a leading cause of such life-threatening infections, being the most frequently isolated gram-negative bacterium from the lower respiratory tract and the second most common cause of bacteremia. At present, there exists no effective immunological means for the control of Klebsiella infections.
Numerous bacterial species, among them Klebsiella, possess a well-defined capsule which surrounds the bacterial cell in a viscous layer. The capsule is composed of repeating units of monosaccharides which form a high molecular weight polymer. Such capsular polysaccharides (CPS) confer the K (or capsular) antigenic specificity upon their respective bacterial cells.
Studies such as described in Riottot, M. M., Fournier, J. M., Pillot, J., "Capsular serotype specificity of the protection conferred on mice by Klebsiella pneumoniae ribosomal preparations," Infect. Immun. 24: 476-482, 1979; Riottot, M. M., Fournier, J. M., Jouin, H., "Direct evidence for the involvement of capsular polysaccharide in the immunoprotective activity of Klebsiella pneumoniae ribosomal preparations," Infect. Immun. 31: 71-77, 1981; and Cooper, J., McA., Rawley, D., "Resistance to Klebsiella pneumoniae and the importance of two bacterial antigens," Austral. J. Expt. Biol. Med. Sci. 60: 629-641, 1982, have shown that an experimental ribosomal vaccine and an experimental killed whole-cell vaccine are capable of providing a degree of protection against Klebsiella infections in experimental animal models. Protection was found to correlate with the induction of a serotype-specific anti-CPS immune response. However, prior attempts to evoke a protective immune response in mice by vaccination with CPS have been heretofore unsuccessful, and this failure has been attributed to the poor immunogenicity of CPS in mice, as noted in J. M. Fournier, C. Jolivet-Reynaud, M. M. Riottot and H. Jouin, Infect. Immun. 32: 420-426, 1981.
It has now been discovered that immunogenic preparations of CPS can be obtained from the culture supernatant of Klebsiella cultivated in a medium designed to support capsule production. These CSP antigens can subsequently be purified to a high degree, and this highly purified product has been shown to be immunogenic and non-pyrogenic in animals, as described in Cryz, S. J., Jr., Furer, E., Germanier, R., "Purification and vaccine potential of Klebsiella capsular polysaccharide," Infect. Immun. 50: 225-280, 1985; and Cryz, S. J., Jr., Furer, E., Germanier, R., "Safety and Immunogenicity of Klebsiella pneumoniae Kl Capsular Polysaccharide Vaccine in Humans," The Journal of Infectious Diseases, Vol. 151, No. 4, April, 1985. Furthermore, as noted in Cryz, S. J., Jr., Furer, E., Germanier, R., "Prevention of fatal experimental burn wound sepsis due to Klebsiella pneumoniae KP1-0 by immunization with homologous capsular polysaccharide," J. Infect. Dis. 150: 817-822, 1984 and Cryz, S. J., Jr., Furer, E. Germanier, R., "Protection against fatal Klebsiella pneumoniae burn wound sepsis by passive transfer of anticapsular polysaccharide," Infect. Immun. 45: 139-142, 1984, anti-serotype-specific CPS has been shown to be highly effective at preventing fatal experimental Klebsiella infections.